Following incubation with insulin, isomethylbutylxanthine (IBMX), and dexamethasone, confluent 3T3-L1 fibroblasts differentiate into cells with morphological and biochemical characteristics of mature rodent adipocytes. Differentiated 3T3-L1 adipocytes and isolated rat fat cells were utilized to investigate the activation of hormone-responsive, particulate cAMP phosphodiesterase (PDE) and the role played by this enzyme in the process of insulin-dependent regulation of lipolysis. In intact 3T3-L1 adipocytes, the antilipolytic agents insulin and phenylisopropyladenosine (PIA) increase particulate cAMP PDE activity. Certain "insulin-like" agents such as wheat germ agglutinin (WGA) and anti-insulin-receptor antibodies increase hexose transport as well as particulate cAMP PDE activity. Effects of PIA, insulin and the "insulin-like" agents on particulate cAMP PDE were prevented in adipocytes exposed to pertussis toxin (PT) suggesting a role for guanyl nucleotide binding proteins in PDE regulation and/or insulin action. With the goal of understanding the molecular regulation of the particulate cAMP PDE by insulin and other agents, lipolysis and particulate cAMP PDE are being studied under identical conditions, i.e., during activation of lipolysis by various combinations of adenosine deaminase and/or isoproterenol (plus/minus adenosine or PIA), and during inhibition of lipolysis by insulin. The particulate-cAMP PDE from rat adipose tissue has been solubilized with polyoxyethylene non-ionic detergents, partially purified and characterized in terms of inhibition by a number of phosphodiesterase inhibitors.